Breckling, B. & Verhoeven, R. (2013) GM-Crop Cultivation – Ecological Effects on a Landscape Scale. Theorie in der Ökologie 17. Frankfurt, Peter Lang.
NOTICE: THIS WORK MAY BE PROTECTED BY COPYRIGHT
YOU ARE REQUIRED TO READ THE COPYRIGHT NOTICE AT THIS LINK BEFORE YOU READ THE FOLLOWING WORK, THAT IS AVAILABLE SOLELY FOR PRIVATE STUDY, SCHOLARSHIP OR RESEARCH PURSUANT TO 17 U.S.C. SECTION 107 AND 108. IN THE EVENT THAT THE LIBRARY DETERMINES THAT UNLAWFUL COPYING OF THIS WORK HAS OCCURRED, THE LIBRARY HAS THE RIGHT TO BLOCK THE I.P. ADDRESS AT WHICH THE UNLAWFUL COPYING APPEARED TO HAVE OCCURRED. THANK YOU FOR RESPECTING THE RIGHTS OF COPYRIGHT OWNERS.
Human cell toxicity of pesticides associated to wide scale agricultural GMOs
Robin Mesnage, Steeve Gress, Nicolas Defarge & Gilles-Eric Séralini
University of Caen, Institute of Biology, CRIIGEN and Risk Pole, MRSH-CNRS, Caen Cedex, France
Agricultural genetically modified (GM) plants are essentially plants which contain pesticides, because they were designed to tolerate or produce pesticides. In 2011, GM crops reached 160 million hectares, with 59 % of herbicide tolerance (mainly Roundup) mostly in soybean, maize, canola, cotton, 15 % of insecticide producing varieties and 26 % combining both traits (James 2011). We characterized cellular side effects of these pesticide residues on non-target human cells. We summarized here our findings:
Glyphosate-based herbicides toxicity
Roundup (R) was highly toxic on human cells, from 10-20 ppm, far below agricultural dilutions. This occurred on hepatic (HepG2, Hep3B and embryonic (HEK293) as well on placental (JEG3) cell lines, but also on human placental extracts, primary umbilical cord cells (HUVEC) and freshly isolated testicular cells (Richard et al. 2005; Benachour et al. 2007; Benachour & Seralini 2009; Gasnier et al. 2010; Clair et al. 2012). All formulations cause total cell death within 24 h, through an inhibition of the mitochondrial succinate dehydrogenase activity, and necrosis, through the release of cytosolic adenylate kinase measuring membrane damage. They also induced apoptosis through the activation of enzymatic caspases 3 / 7 activities. Most importantly, the R commercialized formulation is always more toxic than the active principle alone, the glyphosate (G). These effects were more dependent on the formulation and thus adjuvants content than on the G concentration. We recently measured compositions and effects of 9 Gbased formulations and identified ethoxylated adjuvants (commonly called POEA) as the active principle of cytotoxicity (Messnage et al. 2012a). However, these are considered as inert diluents in international regulations and are not taken into account for chronic effects which are insufficiently tested, and only with G in pre-commercial testing. We previously underlined this loophole (Mesnage 2010). Long term feeding and reproductive trials with pesticides are the only tests long enough to reveal a potential endocrine disruption which was consequently never studied for R until recently (Seralini et al. 2012), however it was for G by itself.
We investigated it by measuring androgen to estrogen conversion by aromatase activity and mRNA on placental human cells and showed that G interacts with the active site of the purified enzyme (Richard et al. 2005). Both parameters were disrupted at subagricultural doses within 24 h. We also observed a human cell endocrine disruption from 0.5 ppm on the androgen receptor in transfected cells, and then from 2 ppm the transcriptional activities on both estrogen receptors which were also inhibited (Gasnier et al. 2009). Aromatase transcription and activity were disrupted from 10 ppm on HepG2. On freshly isolated rat testicular cells, low non-toxic concentrations of R and G (1 ppm) induced a testosterone decrease by 35 % (Clair et al. 2012). This is expected to occur in human cells which are fitted with the same steroidogenic equipment.
G-based formulations are claimed to have been extensively studied by industry and regulatory agencies and are considered as one of the safest pesticides (Williams et al. 2000). This allowed the establishment of high maximum residue limits (MRL) for GM food / feed (up to 400 ppm). For instance, 20 ppm of G are authorized in GM soy and this MRL is in the range of concentrations typically found in a GM soy harvest. In the light of our results, the safety of these thresholds is clearly challenged.
Insecticidal toxins (Bt) toxicity
Modified toxins from Bacillus thuringiensis are Cry proteins forming pores in insect cell membranes (Then 2010). They are claimed and believed to be inert on non-target species. We have tested for the very !rst time Cry1Ab and Cry1Ac modified Bt toxins (10 ppb to 100 ppm) on the HEK293 cell line, as well as their combined actions with R, within 24 h, on three biomarkers of cell death: measurements of mitochondrial succinate dehydrogenase, adenylate kinase release by membrane alterations and caspases 3 / 7 inductions (Mesnage et al. 2012b). Modified Cry1Ab caused cell death from 100 ppm. For Cry1Ac, under such conditions, no effects were detected. In vivo implications should be now assessed, as Cry1Ab does not appear to be proved as an insect specific toxin.
In the new growing generation with stacked traits, G-based herbicides (like R) residues are present in the R-tolerant edible plants and mixed with modified Bt insecticidal toxins that are produced by the GM plants themselves. However, the toxicology of mixtures cannot be fully understood without knowing the combined toxicity of the various compounds of the formulations. In some in vitro conditions, G and its adjuvant synergistically damage cell membranes in a similar manner to R (Benachour & Seralini 2009). R adjuvants change human cell permeability and amplify toxicity induced already by G, through apoptosis and necrosis. The real threshold of G toxicity must take into account the presence of adjuvants but also G metabolism and time-amplified effects or bioaccumulation. For the mixtures of Bt toxins and R, the only measured significant combined effect was that modified Cry1Ab and Cry1Ac reduced caspases 3 / 7 activations induced by R; this could delay the activation of apoptosis and impact on necrosis. There was the same tendency for adenylate kinase activity and succinate dehydrogenase activity measures. Pesticides have to be tested together, 26 % of agricultural GMOs are indeed stacked events.
We also reviewed 19 studies of mammals fed with commercialized GMOs (Seralini et al. 2011). Meta-analysis of all biochemical disruptions indicated liver and kidney problems as end points of GMO diet effects. These are the major reactive organs in case of food chronic intoxication, and several contingent factors suggested that pesticide residues may be involved in the pathological features. All together, our results raise new questions in the risk assessment of food and feed derived from genetically engineered plants.
Benachour, N., Seralini, G.E. (2009) Glyphosate formulations induce apoptosis and necrosis in human umbilical, embryonic, and placental cells. Chem Res Toxicol 22: 97–105.
Benachour, N., Sipahutar, H., Moslemi, S., Gasnier, C., Travert, C., Seralini, G.E. (2007) Time-and dose-dependent effects of roundup on human embryonic and placental cells. Arch Environ Contam Toxicol 53: 126–133.
Clair, E., Mesnage, R., Travert, C., Seralini, G.E. (2012) A glyphosate-based herbicide induces necrosis and apoptosis in mature rat testicular cells in vitro, and testosterone decrease at lower levels. Toxicol In Vitro 26: 269–279.
Gasnier, C., Benachour, N., Clair, E., Travert, C., Langlois, F., Laurant, C., Decroix-Laporte, C., Seralini, G.E. (2010). Dig1 protects against cell death provoked by glyphosate-based herbicides in human liver cell lines. J Occup Med Toxicol 5: 29.
Gasnier, C., Dumont, C., Benachour, N., Clair, E., Chagnon, M.C., Seralini, G.E. (2009) Glyphosate-based herbicides are toxic and endocrine disruptors in human cell lines. Toxicology 262: 184–191.
James, C. (2011) Global Status of Commercialized Biotech/GM Crops: 2011. ISAAA Brief 43. Mesnage, R., Clair, E., Séralini, G.E. (2010) Roundup in genetically modified plants: Regulation and toxicity in mammals. Theorie in der Ökologie 16: 31–33.
Mesnage, R., Bernay, B., Séralini, G.E. (2012a) Ethoxylated adjuvants of glyphosate-based herbicides are active principles of human cell toxicity. Toxicology. doi:10.1016/j.tox.2012.09.006
Mesnage, R., Clair, E., Gress, S., Then, C., Szekacs, A., Seralini, G.E. (2012b) Cytotoxicity on human cells of Cry1Ab and Cry1Ac Bt insecticidal toxins alone or with a glyphosate-based herbicide. J Appl Toxicol. doi: 10.1016/j.tox.2012.09.006
Richard, S., Moslemi, S., Sipahutar, H., Benachour, N., Seralini, G.E. (2005). Differential effects of glyphosate and roundup on human placental cells and aromatase. Environ Health Perspect 113: 716–720.
Seralini, G.E., Mesnage, R., Clair, E., Gress, S., de Vendomois, J., Cellier, D. (2011) Genetically modified crops safety assessments: present limits and possible improvements. Environmental Sciences Europe 23: 10.
Séralini, G.E., Clair, E., Mesnage, R., Gress, S., Defarge, N., Malatesta, M., Hennequin, D., Spiroux de Vendômois, J. (2012) Long term toxicity of a Roundup herbicide and a Roundup-tolerant genetically modified maize. Food Chem Toxicol. 50: 4221–4231.